This proposal will continue the analysis of the mechanism of transformation with the papovavirus, Simian virus 40. We have shown that the stem cell (embryonal carcinoma) of the teratocarcinoma, when infected with SV40 or polyoma virus, does not express viral function (T antigen, V antigen, or infectious virus). This work will be continued by analyzing the presence or absence of the viral DNA, the state of the viral genome, whether the DNA is transcribed, and whether small amounts of viral protein are detectable by techniques other than immunofluorescence. These studies will be useful in defining how the state of differentiation may modulate virus infection and transformation. In the second series of questions, we propose to analyze the virus-cell interactions required for infection and transformation of diploid Chinese hamster cells with SV40 virus. Early after infection, a population (30%) of tetraploid-polyploid cells are induced, which we propose to separate with flow cytometry, and further characterize this population in regard to the acquisition of their malignant phenotype. Further studies are proposed to characterize the mechanism of cell death in populations of SV40 tsA-transformed Chinese hamster cells at the nonpermissive temperature. Various analyses, such as immunoprecipitation of T antigen, analysis of the state of the viral DNA, cell cycle regulation, and certain transformed cell cycle markers are proposed. Finally, we plan to test the tumorigenicity of SV40 tsA-transformed cells in goats (39.5 degrees C temperature), and in the chorioallantoic membrane of the chicken egg. These studies will provide information to and in understanding the mechanism of viral infection and transformation.